QUE-BACT* CRYPTOSPORIDIUM STAINING REAGENTS

Technical Data #8493a / 2013.07.21

The sporozoan parasite, Cryptosporidium species, is considered an overt human pathogen, causing diarrheal disease. Its rate of detection has been about the same as that of any one of the pathogenic Escherichia, Campylobacter, Salmonella, Shigella species and the protozoan Giardia lamblia. (1)(4)

Cryptosporidium oocysts are acid-fast, and this characteristic is used to differentiate them from other non acid-fast structures in fecal specimens.

Of the various acid-fast staining procedures, the dimethyl sulfoxide(DMSO) is the easiest and quickest to perform and provides excellent results. (1) (2)

QUE-BACT* Cryptosporidium staining products are used to stain smears prepared from fecal specimens suspected of containing Cryptosporidium oocysts. DMSO is mixed with the carbol fuchsin stain and transports the carbol fuchsin through the oocyst wall and rapidly stains the red cell without the need to heat the slide. Decolorization and counterstain is done in one step using a mixture of acetic acid and malachite green solution. Oocysts stain red while most other fecal material stains green.

PRODUCTS :

STORAGEWhen not in use, always store in tightly-sealed containers at 15-30ºC in a dark area.

SIGN OF DETERIORATIONStains should not be used if they show signs of deterioration or if the expiration date has passed.

PROCEDURE

Prepare the smear as described in standard references.(5)

Allow the smear on the slide to air dry.

Ensure smear is fixed with QUE BACT crypto Methanol for 5-10 seconds.

Place the fixed smear into the QUE-BACT Crypto Carbol Fuchsin stain for 5 minutes at room temperature.

Wash gently in running tap water, until excess stain is removed.

Place the smear into the QUE-BACT Crypto Malachite Green counterstain decolorizer for 1-2 minutes.

Rinse gently in running tap water for 10 seconds.

Drain and blot dry carefully.

Examine microscopically at 40X.

INTERPRETATIONCryptosporidium oocysts of 4-5 mm.in size should appear as a pink to red color. Other fecal material, including yeasts, will stain green.

LIMITATIONS OF PROCEDUREPositive stains provide presumptive evidence of the presence of Cryptosporidium oocysts. Empty spaces, the size and shape of a Cryptosporidium oocyst may be present in the stained smear. Two sporozoites in an Isospora oocyst, which are larger than a Cryptosporidium oocyst which has four sporozites, may also stain pink to red. Foreign material may stain pink but are different sizes and shapes. A negative staining reaction does not necessarily indicate a negative result. Change staining reagents after 35-40 smears. Other methods may be required.

USER QUALITY CONTROLIt is recommended that control slides be used simultaneously with test slides to increase accuracy.